Lipid abnormalities, such as obesity, hyperlipidemia, and elevated intramyocellular lipid levels have been correlated with the development of insulin resistance, a subnormal metabolic response to insulin. Previous research by our laboratory indicates that ceramide is necessary and sufficient for saturated-fatty-acid-induced insulin resistance in cultured muscle cells. Elevations of ceramide concentration, similar to those seen in cell culture, have been observed in insulin resistant animals and humans. We hypothesize that ceramide is an obligate intermediate in the induction of insulin resistance by saturated fatty acids in vivo. In the project described herein, we will quantitatively determine the role of ceramide in fatty-acid-induced insulin resistance in vivo. Intravenous infusion of saturated and unsaturated lipid emulsions and administration of high fat diets will be used to promote elevations in serum free fatty acids. We will correlate concentrations of total ceramide and individual ceramide species with insulin sensitivity in muscle and liver. Additionally, we will determine whether pharmacological inhibition of ceramide synthesis will prevent the induction of insulin resistance by free fatty acids.